Research Project 2 will bridge the gap between knowledge of the ecology of toxigenic Gambierdiscus on the reef (established in research project 1) and human exposure and toxicity (evaluated in research project 3). Specifically, Project 2 will evaluate the metabolites produced by the toxigenic Gambierdiscus and their flux and mechanisms of transfer into the marine food web to assess the rates of bioaccumulation, mechanisms of transport and biotransformation, and depuration. We will include ecophysiology studies of both toxin and the sulfur osmolyte, dimethylsulfoniopropionate (DMSP) production Gambierdiscus. We hypothesize that DMSP, which is produced at >100 mM intracellular concentrations in Gambierdiscus, may have protective effects (possibly via antioxidant activity) against the effects of the toxin in the producers and consumers as both metabolites move through the food web. The results from these investigations will be critical for the development of toxin flux models and in the assessment of human health risk. Project 2 will also be focused on determining processes/mechanisms that are upregulated in model fish species following acute and sub-lethal exposure, to identify metabolites and biomarkers that may provide insight into the toxicological effects of ciguatoxin. Accordingly, the proposed research will focus on the use of multiple model systems of finfish, exposed to CTX, and use an integrated ?omics? approach, to identify and subsequently validate biomarkers of CTX exposure, as a proxy of likely bioaccumulation. With these results we will move toward development of methods for in situ toxin detection and potential diagnostic or prognostic methodologies. These studies will be coupled with targeted investigations to evaluate cellular CTX transport mechanisms, detoxification mechanisms, DNA damage, and oxidative stress (linked with DMSP production and transformation). The novel integrated approach of project 2, will bring advanced analytical methodologies to bear on a long-standing problem of trying to provide biomarkers and forecasting tools that will ultimately limit exposure of humans to CTX.